We propose to use ISS to develop an improved anthrax vaccine that will be safe and will provide immunity after one or two immunizations. This vaccine will be compromised of recombinant anthrax protective antigen (rPA) delivered with an immunostimulatory (ISS) oligonucleotide adjuvant enhanced by a novel coacervate formulation. We will compare the ISS sequence currently in clinical trials, 1018, with our more active ISS sequence, C274 to identify the most potent vaccine formulation. The use of ISS in the novel coacervate carrier should provide dramatic enhancement of both the speed and magnitude of the antibody response developed against rPA compared to current second generation approaches using rPA adsorbed to alum or compared to the currently licensed vaccine, AVA. The specific aims for this project are: 1. To test the immunogenicity of recombinant anthrax protective antigen (rPA) with and without combination with ISS coacervate formulations in mice, rabbits, and rhesus macaques; sera will be evaluated by PA-specific ELISAs and for the capacity to neutralize the anthrax toxins using in vitro cell culture assays. 2. To establish the ability of 1-2 doses of the optimized ISS coacervate/rPA vaccine to protect experimental animals against anthrax in vivo following challenge with Bacillus anthracis spores. A. Protection of mice against challenge with B. anthracis spores administered by the intranasal route. B. Protection of rabbits against inhalation anthrax following aerosol challenge with B. anthracis spores. C. Protection of rhesus macaques against inhalation anthrax following challenge with B. anthracis spores. 3. To perform toxicological evaluation and to initiate GMP production of the ISS coacervate/rPA vaccine, assuming positive results in Specific Aims 1-3. Because of the high cost of the later Specific Aims (Aims 3C and 4), activities for each Specific Aim will be performed sequentially, and feasibility will be established before the project moves onto the next Aim. Assuming success in the individual Specific Aims, this proposal is intended to produce an IND-ready new anthrax vaccine that could rapidly move into clinical testing.